Expression of shoot-inducing Ti TL-DNA in differentiated tissues of potato (Solanum tuberosum cv Maris Bard)

Abstract

In potato line Mb1501B one or possibly two normal size Ti TL-DNA copies per tetraploid genome were detected by Southern blot analysis, but no TR-DNA. The TL-DNA probably contained the entire transposon Tn1831 inserted at the T-DNA auxin gene for transcript 2. Northern blot analyses of the steady-state RNA in different Mb1501B tissues isolated from (i) shoots cultured in vitro (ii) grafted plants and (iii) tubers, showed that that TL-DNA transcripts 3, 4, 6a and 7 were expressed most abundantly in the cultured shoots. They formed approximately 0.0023 to 0.0007% of the total poly(A) RNA. Transcripts 1, 5 and 6b were not detected in any of the tissues analysed. This indicated even lower levels of expression (below approximately 0.0001% of the total poly(A) RNA or, making certain assumptions, an abundance of less than one T-DNA derived RNA molecule per cell). As expected, transcript 2 was not detected in any of the Mb1501B tissues. The abundance of the transcripts was reduced in grafted plants and tubers compared with cultured shoots with the greatest decrease (5×) for transcripts 4, 6a and 7. Transcript 4, the one most responsible for the changed growth and development of Mb 1501 B, formed approximately 0.0003% of the poly(A) RNA from both grafted plants and tubers.

T-DNA genes to study plant development: precocious tuberisation and enhanced cytokinins in A. tumefaciens transformed potato

“…Careful analysis of tobacco tumours induced by A. tumefaciens mutants inactivated for the gene for transcript 6b suggested that its product might cause a reduction in endogenous auxin levels (29). The abundance of this transcript in MbI501B tissues was below our detection limits (6).…”

“…3c). Confirmation of the transformed status of the tuber-derived MbI501B plant came from further analysis, described in the accompanying paper (6).…”

“…The simple T -D N A structure of M b 1501 B, described in the a c c o m p a n ying paper(6), can be taken as supportive evidence that it contains one type of transformed cells and is likely to have arisen from a single primary transformed cell.Graft ing of line Mb 1501 BThe m o r p h o l o g y of line M b l 5 0 1 B became more like that of Maris Bard when Mb 1501B shoots were grafted repeatedly onto root stocks of untransformed potatoes. Initially, M b I 5 0 1 B shoots were grafted onto rootstocks of in vitro cultured n o r m a l shoots…”

The chloroplast-located glutamine synthetase of Phaseolus vulgaris L.: nucleotide sequence, expression in different organs and uptake into isolated chloroplasts

“…Total RNA was extracted from 10-100 g fresh weight of tissue by the sodium dodecyl sulphate (SDS)-phenol method [5]. Poly(A)+-enriched RNA was purified by two cycles of oligo(dT) cellulose chromatography.…”

Developmental regulation of RI TL-DNA gene expression in roots, shoots and tubers of transformed potato (Solanum tuberosum cv. Desiree)

“…Steady state RNA analysis was done as described previously by Burrell et al (2). Briefly, about 50 g tissue was frozen in liquid nitrogen and RNA extracted in a buffer (200 mM Tris-HC1 pH 8.5; 400 mM NaC1; 60 mM Mg Acetate; 50 mM EGTA; 1%0 SDS; 5 mM DTT and 0.25 M sucrose).…”